In vivo knockdown of gene expression in brain cancer with intravenous RNAi in adutt rats
In vivo knockdown of gene expression in brain cancer with intravenous RNAi in adutt rats
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DOI：
10.1002/jgm.449
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发表时间：
2003-12-01
影响因子：
3.5
通讯作者：
Pardridge, WM
中科院分区：
文献类型：
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作者：
Zhang, Y;Boado, RJ;Pardridge, WM
背景哺乳动物细胞中的基因沉默可以通过RNA干扰（RNAi）与表达质粒进行，所述表达质粒编码短发夹RNA（shRNA），所述短发夹RNA与靶mRNA内的特定序列杂交。在开发RNAi疗法在哺乳动物中的限制因素是基因传递system.Methods本研究描述了生产的抗荧光素酶的shRNA表达质粒，这是封装在85 nm的聚乙二醇化免疫脂质体（PILs）的内部。将荧光素酶基因永久转染的C6大鼠神经胶质瘤细胞植入成年大鼠的尾壳核，其产生表达荧光素酶的颅内脑癌。PIL在体内用针对大鼠转铁蛋白受体（TfR）的单克隆抗体（MAb）靶向穿过血脑屏障和穿过肿瘤细胞膜。TfRMAb系在1-2%的聚（乙二醇）链的尖端共轭的脂质体的表面。结果TfRMAb靶向PILs抑制荧光素酶基因的表达在脑癌的90%，这种效果持续至少5天后，单次静脉注射10 μ g/大鼠的质粒DNA。针对荧光素酶基因的RNAi治疗没有引起肿瘤γ-谷氨酰转肽酶表达的变化。靶向空表达质粒与TfRMAb-PIL导致在脑cancer in vivo.Conclusions荧光素酶活性没有变化，体内RNAi是启用与一种新形式的基因递送系统，封装在PIL的表达质粒，这是有针对性的受体特异性单克隆抗体的基础上的远程站点。PIL基因递送系统和RNAi表达质粒的组合应用使得在静脉内施用后能够在哺乳动物的远端位点（例如脑癌）中进行基因沉默。版权所有（C）2003约翰威利父子有限公司。
Background Gene silencing in mammalian cells is possible with RNA interference (RNAi) with expression plasmids that encode for short hairpin RNAs (shRNA) that hybridize to a specific sequence within a target mRNA. The limiting factor in developing RNAi therapeutics in mammals is the gene delivery system.Methods The present studies describe the production of anti-luciferase shRNA expression plasmids, which are encapsulated in the interior of 85 nm pegylated immunoliposomes (PILs). C6 rat glioma cells, permanently transfected with the luciferase gene, are implanted in the caudate-putamen nucleus of adult rats, which produces luciferase-expressing intracranial brain cancer. The PILs are targeted across the blood-brain barrier and across the tumor cell membrane in vivo with a monoclonal antibody (MAb) to the rat transferrin receptor (TfR). The TfRMAb is tethered to the tips of 1-2% of the poly(ethylene glycol) strands conjugated to the surface of the liposome.Results The TfRMAb-targeted PILs inhibit luciferase gene expression in the brain cancer by 90%, and this effect persists for at least 5 days after a single intravenous injection of 10 mug/rat of plasmid DNA. RNAi therapy directed against the luciferase gene caused no change in expression of tumor gamma-glutamyl transpeptidase. Targeting the empty expression plasmid with the TfRMAb-PIL resulted in no change in luciferase activity in the brain cancer in vivo.Conclusions In vivo RNAi is enabled with a new form of gene delivery system that encapsulates expression plasmids in PILs, which are targeted to distant sites based on the specificity of a receptor-specific monoclonal antibody. The combined application of the PIL gene delivery system and RNAi expression plasmids enables gene silencing in remote sites such as brain cancer in mammals after intravenous administration. Copyright (C) 2003 John Wiley Sons, Ltd.